Recent studies in humans have shown a correlation between advancing age, hypertension, and diabetes and impairment of collateral function/development Controversy currently exists regarding the mechanisms and molecules which mediate "normal" collateral development. Information on molecules and mechanisms responsible for impairment of collateral development is essentially nonexistent. Understanding normal regulatory processes of collateral development and how they are compromised in the presence of major risk factors for vascular disease is a current and long-term goal of this project. Successful remodeling of the vascular wall requires controlled regulation of gene expression, vascular cell growth, and matrix metabolism. Superoxide (02-) and nitric oxide (NO) and their daughter compounds are capable of inhibiting or promoting each of these processes. Studies related to 3 specific aims will determine the role of free radicals in 1) vascular cell growth, 2) gene expression, and 3) matrix metabolism during normal collateral development. Additional experiments within each aim will determine how increased oxidative stress and/or reduced NO bioavailability may impair collateral development. An in vivo model which permits paired comparisons of collateral and control vessels within animals will be used. Spontaneously hypertensive rats (SHR) will be utilized as a model of endothelial dysfunction. Preliminary studies demonstrate that collateral luminal expansion is suppressed in SHR; that inhibition of the NO system mimics this suppression and that antioxidant therapy restores luminal expansion in SHR. In vivo and ex vivo methods will be used to quantify levels of NO and O2-. Specific enzymes, enzyme inhibitors, and free radical scavengers will be used to determine the role of NO and reactive oxygen species. Vascular cell proliferation and apoptosis will be evaluated with immunohistochemical techniques. Gene expression will be determined by real-time RT-PCR and immunoblotting and localized with the arterial wall with immunohistochemistry. Levels of latent and active matrix metaUoproteinase will be determined with gel zymography, immunoblotting and immunohistochemistry. These studies will clarify major mechanisms responsible for normal and impaired collateral growth and predict therapeutic approaches to enhance collateral enlargement.